Composition useful for protein fragment complementation assays (PCA) using fragments of E. coli/TEM-1 β-lactamase

Abstract

The present invention describes an assay method comprising: (A) generating (1) at least a first fragment of a reporter molecule linked to a first interacting domain and at least a second fragment of a reporter molecule linked to a second interacting domain, or (2) nucleic acid molecules that code for (A)(1) and subsequently allowing said nucleic acid molecules to produce their coded products; then, (B) allowing interaction of said domains; and (C) detecting reconstituted reporter molecule activity, where said reporter molecule can react with a penicillin- or a cephalosporin-class substrate.

1. A composition comprising:
 (A) a first compound comprising a first fragment of an interacting domain linked to a first fragment of a β-lactamase reporter enzyme that catalyses the hydrolysis of the amide bond of β-lactam rings in penicillin or cephalosporin compounds; and 
 (B) a second compound comprising a second fragment of an interacting domain linked to a second fragment of said β-lactamase reporter enzyme wherein components (A) and (B) upon complementation results in reconstitution of β-lactamase activity. 
 
     
     
       2. A composition comprising a compound which comprises a fragment of an interacting domain linked to a first fragment of a β-lactamase reporter enzyme that catalyses the hydrolysis of the amide bond of β-lactam rings in penicillin or cephalosporin compounds upon binding to a second complementary fragment of the β-lactamase reporter enzyme. 
     
     
       3. A composition according to  claim 2  or  1  where said interacting domain is derived from a leucine zipper or from a rapamycin-inducible interacting protein. 
     
     
       4. A composition according to  claim 2  or  1  where said interacting domain is derived from a GCN 4 leucine zipper or from FKBP/FRB. 
     
     
       5. A composition according to  claim 2  or  1  wherein at least one of said compounds has a flexible linker joining its reporter molecule fragment to its associated interacting domain.